Bottom Line:
CHMP2B is a component of the ESCRT-III complex, which is required for function of the multivesicular body (MVB), an endosomal structure that fuses with the lysosome to degrade endocytosed proteins.We report a novel endosomal pathology in CHMP2B mutation-positive patient brains and also identify and characterize abnormal endosomes in patient fibroblasts.The fusion of endosomes with lysosomes is required for neuronal function and the data presented therefore suggest a pathogenic mechanism for FTD caused by CHMP2B mutations.

ABSTRACTMutations in CHMP2B cause frontotemporal dementia (FTD) in a large Danish pedigree, which is termed FTD linked to chromosome 3 (FTD-3), and also in an unrelated familial FTD patient. CHMP2B is a component of the ESCRT-III complex, which is required for function of the multivesicular body (MVB), an endosomal structure that fuses with the lysosome to degrade endocytosed proteins. We report a novel endosomal pathology in CHMP2B mutation-positive patient brains and also identify and characterize abnormal endosomes in patient fibroblasts. Functional studies demonstrate a specific disruption of endosome-lysosome fusion but not protein sorting by the MVB. We provide evidence for a mechanism for impaired endosome-lysosome fusion whereby mutant CHMP2B constitutively binds to MVBs and prevents recruitment of proteins necessary for fusion to occur, such as Rab7. The fusion of endosomes with lysosomes is required for neuronal function and the data presented therefore suggest a pathogenic mechanism for FTD caused by CHMP2B mutations.

Mentions:
We have previously shown that overexpression of CHMP2BIntron5 and CHMP2BQ165X produce enlarged late endosomes in human neuroblastoma cells (6). If this endosomal phenotype is relevant to neurodegeneration, we would predict a similar finding in brain tissue. Frontal cortex sections from four FTD-3 patients were immunostained for mannose-6 phosphate receptor (M6PR), a marker of late endosomes. Anti-M6PR labelled large vacuoles in the cytoplasm of cortical neurons which were also visible by haematoxylin and eosin staining (Fig. 2). The vacuolar pathology was most abundant in the deep layers of the cortex. To assess their frequency, we counted the number of neurons that contained enlarged M6PR-positive vacuoles per high power field of cortical layers III to V in three FTD-3 brains. There was a range of 0–15 neurons with vacuolar pathology per high power field (an area of 0.249 mm2) with an average of 2.95 per field (average range 2–4.85). We then determined the distribution of vacuolar pathology in one FTD-3 brain by examining 14 brain regions (Table 1). Vacuoles were found in all cortical areas examined, with the exception of the cerebellar cortex. This is consistent with the generalized cortical atrophy observed on post-mortem (9) and with imaging data showing that atrophy is widespread in the FTD-3 cortex (10,11). The vacuoles were not observed in non-cortical grey matter regions such as the thalamus, substantia nigra, locus coeruleus and medulla. To determine whether these vacuoles were specific to FTD-3, we analysed brains from three Alzheimer's disease patients, six familial FTD patients (four with MAPT mutations, two with GRN mutations), 12 prion disease patients and nine age-matched controls (Supplementary Material, Fig. S1 and Table S1). None of these cases showed enlarged M6PR-positive vacuoles suggesting that this pathology is a result of the CHMP2B mutation.Table 1.

Mentions:
We have previously shown that overexpression of CHMP2BIntron5 and CHMP2BQ165X produce enlarged late endosomes in human neuroblastoma cells (6). If this endosomal phenotype is relevant to neurodegeneration, we would predict a similar finding in brain tissue. Frontal cortex sections from four FTD-3 patients were immunostained for mannose-6 phosphate receptor (M6PR), a marker of late endosomes. Anti-M6PR labelled large vacuoles in the cytoplasm of cortical neurons which were also visible by haematoxylin and eosin staining (Fig. 2). The vacuolar pathology was most abundant in the deep layers of the cortex. To assess their frequency, we counted the number of neurons that contained enlarged M6PR-positive vacuoles per high power field of cortical layers III to V in three FTD-3 brains. There was a range of 0–15 neurons with vacuolar pathology per high power field (an area of 0.249 mm2) with an average of 2.95 per field (average range 2–4.85). We then determined the distribution of vacuolar pathology in one FTD-3 brain by examining 14 brain regions (Table 1). Vacuoles were found in all cortical areas examined, with the exception of the cerebellar cortex. This is consistent with the generalized cortical atrophy observed on post-mortem (9) and with imaging data showing that atrophy is widespread in the FTD-3 cortex (10,11). The vacuoles were not observed in non-cortical grey matter regions such as the thalamus, substantia nigra, locus coeruleus and medulla. To determine whether these vacuoles were specific to FTD-3, we analysed brains from three Alzheimer's disease patients, six familial FTD patients (four with MAPT mutations, two with GRN mutations), 12 prion disease patients and nine age-matched controls (Supplementary Material, Fig. S1 and Table S1). None of these cases showed enlarged M6PR-positive vacuoles suggesting that this pathology is a result of the CHMP2B mutation.Table 1.

Bottom Line:
CHMP2B is a component of the ESCRT-III complex, which is required for function of the multivesicular body (MVB), an endosomal structure that fuses with the lysosome to degrade endocytosed proteins.We report a novel endosomal pathology in CHMP2B mutation-positive patient brains and also identify and characterize abnormal endosomes in patient fibroblasts.The fusion of endosomes with lysosomes is required for neuronal function and the data presented therefore suggest a pathogenic mechanism for FTD caused by CHMP2B mutations.

ABSTRACTMutations in CHMP2B cause frontotemporal dementia (FTD) in a large Danish pedigree, which is termed FTD linked to chromosome 3 (FTD-3), and also in an unrelated familial FTD patient. CHMP2B is a component of the ESCRT-III complex, which is required for function of the multivesicular body (MVB), an endosomal structure that fuses with the lysosome to degrade endocytosed proteins. We report a novel endosomal pathology in CHMP2B mutation-positive patient brains and also identify and characterize abnormal endosomes in patient fibroblasts. Functional studies demonstrate a specific disruption of endosome-lysosome fusion but not protein sorting by the MVB. We provide evidence for a mechanism for impaired endosome-lysosome fusion whereby mutant CHMP2B constitutively binds to MVBs and prevents recruitment of proteins necessary for fusion to occur, such as Rab7. The fusion of endosomes with lysosomes is required for neuronal function and the data presented therefore suggest a pathogenic mechanism for FTD caused by CHMP2B mutations.